[Show abstract][Hide abstract] ABSTRACT: Internal necrosis of carrot has been observed in UK carrots for at least 10 years, and has been anecdotally linked to virus infection. In the 2009 growing season some growers had up to 10% of yield with these symptoms. Traditional diagnostic methods are targeted towards specific pathogens. By using a metagenomic approach with high throughput sequencing technology, other, as yet unidentified causes of root necrosis were investigated. Additionally a statistical analysis has shown which viruses are most closely associated with disease symptoms. Carrot samples were collected from a crop exhibiting root necrosis (102 Affected: 99 Unaffected) and tested for the presence of the established carrot viruses: Carrot red leaf virus (CtRLV), Carrot mottle virus (CMoV), Carrot red leaf associated viral RNA (CtRLVaRNA) and Parsnip yellow fleck virus (PYFV). The presence of these viruses was not associated with symptomatic carrot roots either as single viruses or in combinations. A sub-sample of carrots of mixed symptom status was subjected to MiSeq sequencing. The results from these tests suggested Carrot yellow leaf virus (CYLV) was associated with symptomatic roots. Additionally a novel Torradovirus, a novel Closterovirus and two novel Betaflexiviradae related plant viruses were detected. A specific diagnostic test was designed for CYLV. Of the 102 affected carrots, 98% were positive for CYLV compared to 22% of the unaffected carrots. From these data we conclude that although we have yet to practically demonstrate a causal link, CYLV appears to be strongly associated with the presence of necrosis of carrots.
[Show abstract][Hide abstract] ABSTRACT: The 9573-nucleotide genome of a potyvirus was sequenced from a Coriandrum sativum plant from India with viral symptoms. On analysis, this virus was shown to have greater than 85 % nucleotide sequence identity to vanilla distortion mosaic virus (VDMV). Analysis of the putative coat protein sequence confirmed that this virus was in fact VDMV, with greater than 91 % amino acid sequence identity. The genome appears to encode a 3083-amino-acid polyprotein potentially cleaved into the 10 mature proteins expected in potyviruses. Phylogenetic analysis confirmed that VDMV is a distinct but ungrouped member of the genus Potyvirus.
[Show abstract][Hide abstract] ABSTRACT: Xanthomonas campestris pathovar musacearum (Xcm) is the causal agent of banana Xanthomonas wilt, a major threat to banana production in eastern and central Africa. The pathogen is present in very high levels within infected plants and can be transmitted by a broad range of mechanisms; therefore early specific detection is vital for effective disease management. In this study we have developed a polyclonal antibody (pAb) and deployed this in a lateral flow device (LFD) format to allow rapid in-field detection of Xcm. We also independently assessed published Xcm PCR assays: only two assays gave specific amplification of Xcm, whilst others cross-reacted with non-target Xanthomonas species. Pure cultures of Xcm were used to immunise a rabbit, the IgG antibodies purified from the serum and the resulting polyclonal antibodies tested using ELISA and LFD. Testing against a wide range of bacterial species showed the pAb detected all strains of Xcm, representing isolates from seven countries and the known genetic diversity of Xcm. The pAb also detected the closely related Xanthomonas axonopodis pathovar vasculorum (Xav), primarily a sugarcane pathogen. Detection was successful in both naturally and experimentally infected banana plants, and the LFD limit of detection was 105 cells/ml. Whilst the pAb is not fully specific for Xcm, Xav has never been found in banana. Therefore the LFD can be used as a first line screening tool to detect Xcm in the field. Testing by LFD requires no equipment, can be performed by non-scientists and is cost-effective. Therefore this LFD provides a vital tool to aid in the management and control of Xcm.This article is protected by copyright. All rights reserved.
[Show abstract][Hide abstract] ABSTRACT: Maize lethal necrosis (MLN) is a disease of maize caused by the combination of Maize chlorotic mottle virus (MCMV) and a potyvirus (Uyemote et al., 1981). MLN caused by MCMV and the potyvirus Sugarcane mosaic virus (SCMV) was identified in Kenya in 2012 (Adams et al., 2013) and associated with significant damage. Subsequent to the outbreak in Kenya, MLN symptoms have been associated with MCMV and SCMV in Tanzania and Uganda and, based on our initial findings, MCMV but not SCMV in Rwanda (ASARECA, 2013). The control of MLN is a priority for the East Africa region. We report here a fuller investigation of the causal agents of MLN from Rwanda.
In March and June 2013 samples of maize showing signs of MLN symptoms were received from the Northern Province of Rwanda for MCMV and SCMV testing. The samples were tested by real-time PCR as described in Adams et al. (2013). By this method all samples tested positive for MCMV and negative for SCMV. In order to determine if MCMV was responsible for the symptoms, or if another potyvirus was present, deep sequencing of total RNA extracted from four of the maize samples was performed. Indexed ScriptSeq libraries (Epibio, UK) were produced and sequenced on a MiSeq sequencer (Illumina, UK) using 500 cycle V2 reagents. The resulting 10 million 250 bp paired-end reads were split by index, quality filtered to a score of above Q20 and assembled using Trinity (Grabherr et al., 2011). Contigs over 200 bp were then compared to the GenBank nr database using BLASTX and the resulting data analysed using MEGAN as previously reported to generate draft genomes (Adams et al., 2013).
Complete genomes of MCMV were recovered from all four maize samples from Rwanda (GenBank Accession Nos. KF744393-KF744396) and shown to have a very close relatedness with MCMV from Kenya and China (99% homology), and with some separation from MCMV from the United States (97-96% homology) (Fig. 1). Complete SCMV genomes were found in three of the four samples (KF744390- KF744392). Fig. 2 shows the relationship between these and previous isolates. The SCMV isolated from Rwandan samples is distinct from that isolated in Kenya (87% identity) and most closely related (95% identity) to strain SCMV-DMB (BA00797). A closer examination of the sequence at the primer binding regions of the respective SCMV genomes shows a high degree of divergence and explain the negative results following real-time PCR testing using the assay designed to the SCMV sequence from Kenya (Adams et al., 2013).
These data would support a shared origin for MCMV in Rwanda and Kenya, but different sources for SCMV. This is not unexpected as SCMV has been observed for many years in East Africa (Louie, 1980), whereas MCMV is believed to be new to the region (Adams et al., 2013). MCMV would now appear to be spreading across the East Africa region, potentially in seed, and causative of MLN where populations of SCMV are already endemic. These findings have implications for the spread of MLN throughout Africa and for the testing for MCMV and SCMV. Control of the disease may prove more effective by initiating quarantine measures to counter the movement of MCMV, rather than attempting to control the endemic SCMV. This report confirms that MLN, caused by MCMV and SCMV, is now present in Rwanda and that this is most probably due to the recent introduction of MCMV.
[Show abstract][Hide abstract] ABSTRACT: In Europe the most devastating phytoplasma associated with grapevine yellows (GY) diseases is a quarantine pest flavescence dorée (FDp) from the 16SrV taxonomic group. The on-site detection of FDp with an affordable device would contribute to faster and more efficient decisions on the control measures for FDp. Therefore, a real-time isothermal LAMP assay for detection of FDp was validated according to the EPPO standards and MIQE guidelines. The FD LAMP assay was shown to be specific and extremely sensitive, since it detected FDp in all leaf samples that were determined to be FDp infected using quantitative real-time PCR. The whole procedure of sample preparation and testing was designed and optimised for on-site detection and can be completed in one hour. The homogenisation procedure of the grapevine samples (leaf vein, flower or berry) was optimised to allow direct testing of the crude homogenates with the LAMP assay, without the need for a DNA extraction, and was shown to be extremely sensitive.This article is protected by copyright. All rights reserved.
[Show abstract][Hide abstract] ABSTRACT: Partial recombinant secA proteins were produced from six different phytoplasma isolates representing five 16Sr groups and the expressed, purified recombinant (partial secA) protein from Cape St. Paul wilt disease phytoplasma (CSPWD, 16SrXXII) was used to immunise mice. Monoclonal antibodies (mAbs) were selected by screening hybridoma supernatants for binding to the recombinant proteins. To characterise the binding to proteins from different phytoplasmas, the antibodies were screened by ELISA and western blotting, and epitope mapping was undertaken. Eight different mAbs with varying degrees of specificity against recombinant proteins from different phytoplasma groups were selected. Western blotting revealed that the mAbs bind to proteins in infected plant material, two of which were specific for phytoplasmas. ELISA testing of infected material, however, gave negative results suggesting that either secA was not expressed at sufficiently high levels, or conformational changes of the reagents adversely affected detection. This work has shown that the phytoplasma secA gene is not a suitable antibody target for routine detection, but has illustrated proof of principle for the methodology.
[Show abstract][Hide abstract] ABSTRACT: The performance of probes on an oligonucleotide microarray can be characterised in terms of hybridisation signal strength and the ability to discriminate sequence mismatches between the probe and the hybridising target strand, such as those resulting from SNPs. Various properties of the probe affect mismatch discrimination, such as probe length and the position of mismatched bases, and the effects of these factors have been well characterised in a variety of array formats.
A low-density microarray was developed to systematically investigate the effect of a probe's position within hybridised target PCR products on the tolerance and discrimination of single-nucleotide mismatches between the probe and target. In line with previous reports, hybridisation signals were attenuated by different degrees depending on the identity of the mismatch, the position of the mismatch within the probe, and the length of the PCR product. However, the same mismatch caused different degrees of attenuation depending on the position of the probe within the hybridising product, such that improved mismatch discrimination was observed for PCR products where a greater proportion of the total length was proximal to the array surface.
These results suggest that the degree of mismatch discrimination can be influenced by the choice of PCR primers, providing a means by which array performance could be fine-tuned in addition to manipulation of the properties of the probes themselves.
[Show abstract][Hide abstract] ABSTRACT: Despite the seemingly continuous development of newer and ever more elaborate methods for detecting and identifying viruses, very few of these new methods get adopted for routine use in testing laboratories, often despite the many and varied claimed advantages they possess. To understand why the rate of uptake of new technologies is so low, requires a strong understanding of what makes a good routine diagnostic tool to begin. This can be done by looking at the two most successfully established plant virus detection methods: enzyme-linked immunosorbant assay (ELISA) and more recently introduced real-time polymerase chain reaction (PCR). By examining the characteristics of this pair of technologies, it becomes clear that they share many benefits, such as an industry standard format and high levels of repeatability and reproducibility. These combine to make methods that are accessible to testing labs, which are easy to establish and robust in their use, even with new and inexperienced users. Hence, to ensure the establishment of new techniques it is necessary to not only provide benefits not found with ELISA or real-time PCR, but also to provide a platform that is easy to establish and use. In plant virus diagnostics, recent developments can be clustered into three core areas: 1. techniques that can be performed in the field or resource poor locations (e.g., loop-mediated isothermal amplification LAMP); 2. multiplex methods that are able to detect many viruses in a single test (e.g., Luminex bead arrays); and 3. methods suited to virus discovery (e.g., next generation sequencing, NGS). Field based methods are not new, with Lateral Flow Devices (LFDs) for the detection being available for a number of years now. However the widespread uptake of this technology remains poor. LAMP does offer significant advantages over LFDs, in terms of sensitivity and generic application, but still faces challenges in terms of establishment. It is likely that the main barrier to the uptake of field-based technologies is behavioral influences, rather than specific concerns about the performance of the technologies themselves. To overcome this, a new relationship will need to develop between centralized testing laboratories offering services and those requiring tests; a relationship which is currently in its infancy. Looking further into the future, virus discovery and multiplex methods seem to converge as NGS becomes ever cheaper, easier to perform and can provide high levels of multiplexing without the use of virus specific reagents. So ultimately the key challenge from a routine testing lab perspective will not be one of investment in platforms-which could even be outsourced to commercial sequencing services-but one of having the skills and expertise to analyse the large datasets generated and their subsequent interpretation. In conclusion, only time will tell which of the next-generation of methods currently in development will become the routine diagnostics of the future. This will be determined through a combination of factors. And while the technology itself will have to offer performance advantages over existing methods in order to supplant them, it is likely to be human factors e.g., the behaviors of end users, laboratories and policy makers, the availability of appropriate expertise, that ultimately determine which ones become established. Hence factors cannot be ignored and early engagement with diagnostic stakeholders is essential.
[Show abstract][Hide abstract] ABSTRACT: The complete nucleotide sequences of RNA 1 and RNA 2 of the nepovirus potato black ringspot virus (PBRSV) from two different isolates were determined, as well as partial sequences from two additional isolates. RNA1 is 7,579-7,598 nucleotides long and contains one single open reading frame (ORF), which is translated into a large polyprotein with 2,325 amino acids and a molecular weight of 257 kDa. The complete sequence of RNA2 ranges from 3857 to 3918 nt between the different isolates. It encodes a polyprotein of 1079-1082 amino acids with a molecular weight of 120 kDa. Sequence comparison using the Pro-Pol region and CP showed that all four isolates formed two distinct groups, corresponding to potato and arracacha, that were closely related to each other and also to tobacco ringspot virus (TRSV). Comparing our data to those obtained with other nepoviruses, our results confirm that PBRSV belongs to a distinct species and is a member of subgroup A in the genus Nepovirus based on its RNA2 size, genome organization, and nucleotide sequence.
Archives of Virology 10/2013; · 2.28 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: This study reports the first complete genome sequence of Piper yellow mottle virus (PYMoV, KC808712) identified in black pepper. The genome is 7,622 nucleotides long, possessing four open reading frames (ORFs). ORF1, ORF2 and ORF4 of PYMoV are reported as hypothetical proteins of unknown function with a predicted molecular mass of 15.7, 17.1 and 17.9 kDa, respectively. ORF3 of PYMoV encodes a polyprotein of 218.6 kDa and consists of a viral movement protein (MP), trimeric dUTPase, zinc finger, retropepsin, RT-LTR, and RNAse H. Detailed PYMoV genome analysis confirmed that it is a member of the family Caulimoviridae, genus Badnavirus. Fragments of two additional novel sequences resembling those found in members of the family Caulimoviridae were also identified in the black pepper sample, and the viruses from which they were derived were tentatively named Piper DNA virus 1 and 2.
Archives of Virology 09/2013; · 2.28 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: The availability of characterised reference isolates of plant pathogens is of crucial importance for research and diagnostic laboratories. The Q-bank Plant Viruses and Viroids database contains data and information on plant viruses and viroids, with the unique feature that it is linked to specimens present in publicly available physical collections. The Q-bank database aims to share data and information on the virus and viroid species and their availability between research and diagnostic laboratories. Currently the database focuses on regulated virus species. In future more plant viruses and viroids will be included to provide a comprehensive data and information system. The curators invite virologists worldwide to participate in this international initiative by making their data and isolates available via Q-bank (http://www.q-bank.eu). La base de données et les collections Virus et Viroïdes des Plantes de Q-bank La disponibilité d'isolats de référence caractérisés est d'une importance cruciale pour les laboratoires de recherche et de diagnostic. La base de données de Q-bank sur les Virus et Viroïdes des plantes contient des données et des informations sur les virus et viroïdes, avec l'avantage d’être liée à des spécimens présents dans des collections physiques qui sont disponibles publiquement. La base de données Q-bank vise à permettre le partage de données et d'informations sur les espèces de virus et de viroïdes et leur mise à disposition entre laboratoires de recherche et de diagnostic. Actuellement, la base de données se concentre sur les espèces de virus réglementées. A l'avenir, d'autres virus et viroïdes des végétaux seront ajoutés pour fournir un système de données et d'informations cohérent et complet. Les curateurs invitent les virologistes dans le monde entier à participer à cette initiative internationale en rendant leurs données et leurs isolats disponibles au travers de Q-bank (http://www.q-bank.eu). Бaзa дaнныч о вирусaч и вироидaч рaстений и коллекции Q-bank Для иccлeдoвaтeльcкиx и диaгнocтичecкиx лaбopaтopий дocтyпнocть oxapaктepизoвaнныx cпpaвoчныx изoлятoв пaтoгeнoв pacтeний имeeт пepвocтeпeннoe знaчeниe. Бaзa дaнныx пoд нaзвaниeм «The Q-bank Plant Viruses & Viroids» coдepжит дaнныe и инфopмaцию o виpycax и виpoидax pacтeний и имeeт тy yникaльнyю ocoбeннocть, чтo oнa cвязaнa c экзeмпляpaми, нaличecтвyющими в пyбличнo дocтyпныx кoллeкцияx. Бaзa дaнныx Q-bank cтaвит cвoeй цeлью oбмeн дaнными и инфopмaциeй o видax виpycoв и виpoидax и иx дocтyпнocти для иccлeдoвaтeльcкиx и диaгнocтичecкиx лaбopaтopий. B нacтoящee вpeмя бaзa дaнныx cocpeдoтoчeнa нa peгyлиpyeмыx видax виpycoв. B бyдyщeм в нee бyдeт включeнo бoльшee кoличecтвo виpycoв и виpoидoв pacтeний, c тeм чтoбы oбecпeчивaлacь пoлнoтa дaнныx и инфopмaциoнныx cиcтeм. Кypaтopы этoй бaзы пpeдлaгaют виpoлoгaм вceгo миpa пoyчacтвoвaть в этoй мeждyнapoднoй инициaтивe пyтeм пpeдocтaвлeния cвoиx дaнныx и изoлятoв чepeз Q-bank (<http://www.q-bank.eu>).
[Show abstract][Hide abstract] ABSTRACT: Rhizoctonia solani is an important pathogen of potatoes causing stem canker and black scurf. The fungus is a species complex comprised of 13 known anastomosis groups (AGs). AG3-PT is the anastomosis group frequently associated with disease in potatoes. A real-time PCR assay was designed to the rDNA ITS region of AG3-PT isolates to enable the pathogen to be detected directly in tuber and soil samples. The resulting assay was highly specific for AG3-PT, and did not amplify DNA from isolates from other AGs or subgroups of AG3. Using a bulk DNA extraction method capable of extracting from up to 250 g of soil, the assay could detect one individual sclerotium of AG3-PT (weighing 200 μg) in 250 g of soil. The AG3-PT assay was used, with assays for AG2-1, AG5 and AG8 to determine the prevalence of those AGs in UK potato soils and tubers. AG2-1 and AG3-PT were the predominant groups in tubers and soils, although AG3-PT was more frequently isolated from tubers, highlighting its importance as a potato pathogen. AG3-PT was also detected in more than half of the tuber samples tested suggesting the importance of seed borne inoculum.
European Journal of Plant Pathology 06/2013; 136(2). · 1.71 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: Loop-mediated isothermal AMPlification (LAMP) is an alternative amplification technology which is highly sensitive and less time consuming than conventional PCR-based methods. Three LAMP assays were developed, two for detection of species of symbiotic blue-stain fungi associated with Ips acuminatus, a bark beetle infesting Scots pine (Pinus sylvestris), and an additional assay specific to I. acuminatus itself for use as a control. In common with most bark beetles, I. acuminatus is associated with phytopathogenic blue-stain fungi involved in the process of exhausting tree defenses, which is a necessary step for the colonization of the plant by the insect. However, the identity of the main blue-stain fungus vectored by I. acuminatus was still uncertain, as well as its frequency of association with I. acuminatus under outbreak and non-outbreak conditions. In this study, we employed LAMP technology to survey six populations of I. acuminatus sampled from the Southern Alps. Ophiostoma clavatum was detected at all sampling sites, while O. brunneo-ciliatum, reported in part of the literature as the main blue-stain fungus associated with I. acuminatus, was not detected on any of the samples. These results are consistent with the hypothesis that O. clavatum is the main blue-stain fungus associated with I. acuminatus in the Southern Alps. The method developed in the course of this work provides a molecular tool by which it will be easy to screen populations and derive important data regarding the ecology of the species involved.
Applied and Environmental Microbiology 02/2013; · 3.95 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: Guignardia citricarpa Kiely (anamorph Phyllosticta citricarpa Van der Aa), the causal agent of citrus black spot disease, is subject to phytosanitary restrictions in the EU and USA, such that consignments of citrus are rejected at import if citrus black spot is identified on inspection. Due to the variability of black spot symptoms, positive identification solely on the basis of visual inspection is difficult, especially when lesions lack pycnidia (fruiting bodies of the anamorph Phyllosticta citricarpa). As an aid to visual inspection of symptoms, we have developed a method for detection of G. citricarpa using loop-mediated isothermal amplification (LAMP) which can be used to confirm the presence of G. citricarpa in black spot lesions, including those lacking pycnidia. The LAMP assay can be used to test crude extracts prepared directly from lesions on fruit, and the entire test can be completed in less than 40 min, making it faster than previously described PCR-based methods for detection of G. citricarpa. The method is sufficiently simple to allow deployment of the test in the field, for example in the course of import inspections. Recent years have seen the description of a number of newly recognised species in the genus Phyllosticta that are associated with citrus. As new species emerge, and the taxonomy of the genus is resolved, it will be important to periodically re-evaluate the performance of DNA-based methods for detection of G. citricarpa, including the LAMP assay described here, such that the accuracy of diagnosis can be assured.
European Journal of Plant Pathology 01/2013; 136(2). · 1.71 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: The objective of this study was to develop a rapid, sensitive detection assay for the quarantine pathogen Xanthomonas arboricola pv. pruni, causal agent of stone fruit bacterial spot, an economically important disease of Prunus spp. Unique targets were identified from X. arboricola pv. pruni genomes using a comparative genomics pipeline of other Xanthomonas species, subspecies and pathovars, and used to identify specific diagnostic markers. Loop-mediated isothermal amplification (LAMP) was then applied to these markers to provide rapid, sensitive and specific detection. The method developed showed unrivalled specificity with the 79 tested strains and, in contrast to previously established techniques, distinguished between phylogenetically close subspecies such as X. arboricola pv. corylina. The sensitivity of this test is comparable to that of a previously reported TaqMan™ assay at 103 CFU mL−1, while the unrivalled speed of LAMP technology enables a positive result to be obtained in
Journal of Microbiological Methods 01/2013; 92:332-339. · 2.16 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: Several molecular methods have been developed for detection of Erwinia amylovora, the causal agent of fire blight in pear and apple, but none are truly applicable for on-site use in the field. We developed a fast, reliable and field applicable detection method using a novel target on the E. amylovora chromosome that we identified by applying a comparative genomics pipeline. The target coding sequences (CDS) are both uniquely specific for and all-inclusive of E. amylovora genotypes. This avoids potential false negatives that can occur with most commonly used methods based on amplification of plasmid gene targets, which can vary among strains. Loop-mediated isothermal AMPlification (LAMP) with Optigene GenieII chemistry and instrumentation proved to be an exceptionally rapid (under 15 minutes) and robust method for detecting E. amylovora in orchards, as well as simple to use in the plant diagnostic laboratory. Comparative validation results using plant samples from inoculated greenhouse trials and from natural field infections (of regional and temporal diverse origin) showed that our LAMP had equivalent or greater performance regarding sensitivity, specificity, speed and simplicity than real-time PCR (TaqMan), other LAMP assays, immunoassays and plating, demonstrating its utility for routine testing.
Journal of microbiological methods 12/2012; · 2.43 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: The complete genome sequences of RNA1 and RNA2 of the oca strain of the potato virus arracacha virus B were determined using next-generation sequencing. The RNA1 molecule is predicted to encode a 259-kDa polyprotein with homology to proteins of the cheraviruses apple latent spherical virus (ALSV) and cherry rasp leaf virus (CRLV). The RNA2 molecule is predicted to encode a 102-kDa polyprotein which also has homology to the corresponding protein of ALSV and, to a lesser degree, CRLV (30 % for RNA1, 24 % for RNA2). Detailed analysis of the genome sequence confirms that AVB is a distinct member of the genus Cheravirus.
Archives of Virology 11/2012; · 2.28 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: A real-time PCR assay specific for Sclerotium cepivorum, the causal agent of white rot in onions, was developed for use with a new DNA extraction method capable of processing up to 1 kg of soil in weight. The assay was specific for S. cepivorum when tested against 24 isolates representative of 14 closely related species and other pathogens of onion. The assay was highly sensitive when used with soil DNA extracted using the new DNA extraction procedure. In three different field soils tested, a good relationship between cycle threshold (Ct) and number of sclerotia was observed (R² = 0.89). Twenty-nine soil samples from onion and leek crops were obtained and the pathogen was detected in four samples. All four positive samples were associated with current or past outbreaks of white rot of onion. Additional assays were also used on the 29 field soil samples, Botrytis aclada and Rhizoctonia solani AG8 were also detected, in one soil sample each. Rhizoctonia solani AG2-1 was more widespread and was detected in eight different soil samples. The method is therefore suitable to quantify levels of S. cepivorum in soil samples, with the added advantage that the technique allows other soil pathogens of interest to be assayed from the same DNA sample. The bulk soil DNA extraction method described here has the potential to be used to detect soil-borne pests and pathogens for other crops in a wide range of soil types.
European Journal of Plant Pathology 11/2012; 143(3):467-473. · 1.71 Impact Factor